技术资料

Much attention is focused on characterizing the contribution of bone marrow (BM)-derived cells to regenerating skeletal muscle,fuelled by hopes for stem cell-mediated therapy of muscle degenerative diseases. Though physical integration of BM stem cells has been well documented,little evidence of functional commitment to myotube phenotype has been reported. This is due to the innate difficulty in distinguishing gene products derived from donor versus host nuclei. Here,we demonstrate that BM-derived stem cells contribute via gene expression following incorporation to skeletal myotubes. By co-culturing human BM-derived mesenchymal stem cells (MSC) with mouse skeletal myoblasts,physical incorporation was observed by genetic lineage tracing and species-specific immunofluorescence. We used a human-specific antibody against the intermediate filament protein nestin,a marker of regenerating skeletal muscle,to identify functional contribution of MSC to myotube formation. Although nestin expression was never detected in MSC,human-specific expression was detected in myotubes that also contained MSC-derived nuclei. This induction of gene expression following myotube integration suggests that bone marrow-derived stem cells can reprogram and functionally contribute to the muscle cell phenotype. We propose that this model of myogenic commitment may provide the means to further characterize functional reprogramming of MSC to skeletal muscle. View Publication

Adult hepatocytes are polarised with their apical and basolateral membranes separated from neighbouring cells by tight junction proteins. Although efficient differentiation of pluripotent stem cells to hepatocytes has been achieved,the formation of proper polarisation in these cells has not been thoroughly investigated. In the present study,human embryonic stem cells (hESCs) and human mesenchymal stem cells (hMSCs) were differentiated to hepatocyte-like cells and the derived hepatocytes were characterised for mature hepatocyte markers. The secretion of hepatic proteins,expression of hepatic genes and the functional hepatic polarisation of stem cell-derived hepatocytes,foetal hepatocytes and the HepG2 hepatic cell line were evaluated and the different lines were compared. The results indicate that hESC-derived hepatocytes are phenotypically more robust and functionally more efficient compared with the hMSC-derived hepatocytes,suggesting their suitability for toxicity studies. View Publication

Current therapies for delayed- or nonunion bone fractures are still largely ineffective. Previous studies indicated that the VEGF homolog placental growth factor (PlGF) has a more significant role in disease than in health. Therefore we investigated the role of PlGF in a model of semi-stabilized bone fracture healing. Fracture repair in mice lacking PlGF was impaired and characterized by a massive accumulation of cartilage in the callus,reminiscent of delayed- or nonunion fractures. PlGF was required for the early recruitment of inflammatory cells and the vascularization of the fracture wound. Interestingly,however,PlGF also played a role in the subsequent stages of the repair process. Indeed in vivo and in vitro findings indicated that PlGF induced the proliferation and osteogenic differentiation of mesenchymal progenitors and stimulated cartilage turnover by particular MMPs. Later in the process,PlGF was required for the remodeling of the newly formed bone by stimulating osteoclast differentiation. As PlGF expression was increased throughout the process of bone repair and all the important cell types involved expressed its receptor VEGFR-1,the present data suggest that PlGF is required for mediating and coordinating the key aspects of fracture repair. Therefore PlGF may potentially offer therapeutic advantages for fracture repair. View Publication

Hematopoiesis is maintained by specific interactions between both hematopoietic and nonhematopoietic cells. Whereas hematopoietic stem cells (HSCs) have been extensively studied both in vitro and in vivo,little is known about the in vivo characteristics of stem cells of the nonhematopoietic component,known as mesenchymal stem cells (MSCs). Here we have visualized and characterized human MSCs in vivo following intramedullary transplantation of enhanced green fluorescent protein-marked human MSCs (eGFP-MSCs) into the bone marrow (BM) of nonobese diabetic/severe combined immunodeficiency (NOD/SCID) mice. Between 4 to 10 weeks after transplantation,eGFP-MSCs that engrafted in murine BM integrated into the hematopoietic microenvironment (HME) of the host mouse. They differentiated into pericytes,myofibroblasts,BM stromal cells,osteocytes in bone,bone-lining osteoblasts,and endothelial cells,which constituted the functional components of the BM HME. The presence of human MSCs in murine BM resulted in an increase in functionally and phenotypically primitive human hematopoietic cells. Human MSC-derived cells that reconstituted the HME appeared to contribute to the maintenance of human hematopoiesis by actively interacting with primitive human hematopoietic cells. View Publication